Genital herpes is one of the most prevalent sexually transmitted infections worldwide and its synergistic relationship with HIV amplifies its public health implications. Oral suppressive acyclovir failed to reduce the risk of HIV acquisition or prevent transmission of HIV from HIV/HSV-2 co-infected individuals to their HIV-naive partners highlighting the need to further define the biological mechanisms by which these two pathogens interact. Studies supported by this R01 identified several potential mechanisms, focusing on interactions between HSV and mucosal immunity. Results demonstrate that female genital tract secretions inhibit HIV and HSV in vitro, reflecting the activities of antimicrobial peptides. However, HSV interferes with this defense by downregulating protective mediators, which could lead to a loss in the anti-HIV activity of genital tract secretions. In addition, immature dendritic cells respond to HSV by releasing inflammatory cytokines and chemokines sufficient to increase HIV replication, but paradoxically, fail to mature. The HSV infected and uninfected bystander dendritic cells are rendered incapable of responding to subsequent antigenic stimuli. Thus, in the presence of overt or subclinical HSV infection, mucosal dendritic cells would be incapable of presenting HIV peptides to T cells. HSV also interferes with anatomic host defenses. HSV disrupts epithelial cell tight junctions, greatly facilitating migration of HIV across the epithelial barrier. These results complement the clinical observation that activated T cells persist at sites of HSV-2 reactivation for months after healing of genital ulcers. Together, these findings support the overall paradigm that HSV renders the genital tract mucosal environment more permissive for HIV acquisition and transmission. To test this paradigm with the overall goal of identifying novel strategies to disrupt the synergy between HSV and HIV, we will expand the humanized bone marrow-liver-thymus (BLT) mouse model of HIV to a model of HIV-1/HSV-2 mucosal co- infection and examine the impact HSV has on mucosal immunity, investigate how these changes promote HIV acquisition and increase HIV replication, and determine whether sustained acyclovir delivery using implants similar to intravaginal ring devices will be more effective than intermittent acyclovir dosing at blocking the mucosal responses to HSV that promote HIV infection. In parallel with the proposed mouse studies, we will conduct a prospective study among HIV-1/HSV-2 co-infected and HIV-1?2- women. We will use self- collected vaginal swabs to measure the impact of HSV reactivation on endogenous anti-HIV and anti-HSV activity in genital tract secretions and correlate results with genital tract HIV and HSV viral loads. Additional clinical samples (cytobrush and biopsy) will be collected during episodes of clinical reactivation to assess changes in immune cell populations in response to viral reactivation.